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I. M. Asota, R. Stulting, B. Fant, H. F. Edelhauser; The Effect of Ultraviolet-Light-A Crosslinking on Corneal Stroma Riboflavin Uptake. Invest. Ophthalmol. Vis. Sci. 2008;49(13):3914.
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© ARVO (1962-2015); The Authors (2016-present)
To define optimal levels of 0.1% riboflavin-20% dextran in rabbit corneas to be used for crosslinking with ultraviolet-light-A in humans with keratoconus.
Four pretreatment parameters were tested on albino rabbit globes (Pel-Freez) without the epithelium corresponding to current protocols used in humans for riboflavin-UVA-cross-linking: 20ul of 0.1% riboflavin-20% dextran (Medio-Cross) every minute for five and ten min, and every two and five min for 30 minutes. After pretreatment, the corneas were either treated with UVA (365+/-10 nm, UV-XTM) for up to 30 min or left untreated. Each experiment (N=30) was stopped at 5- to 10-min intervals up to an hour after treatment. Corneal buttons (8mm) were excised and the concentration of riboflavin (24-hour washout in 1 ml of deionized water) was measured with a fluorometer (TD-700). Frozen sections of the corneas were obtained corresponding to the time after pretreatment and UV exposure.
There was little diffusion of riboflavin into the corneal stroma with intact epithelium, with or without UVA. Riboflavin concentrations (mean +/- SD) in deepithelialized corneas treated every 1 min for 5 min were the lowest, peaking at 10 min after treatment at 0.42+/-0.06 mcg/mL and decreasing after 20 min.; and, highest with every 5 min for 30 min., peaking at 0.58+/-0.04 mcg/mLg/ml. Pretreating every 1 min. for 10 min. produced similar concentrations to every 5 min for 30 min., peaking at 0.52 +/- 0.076mcg/mL. Peak concentrations were achieved within 10 min after completion of the pretreatment in the absence of UVA light and slowly decreasing thereafter. UVA light markedly decreased stromal riboflavin concentration for all pretreatment regimens, with peak concentrations reached at 5 min after completion of the pretreatment and the start of UVA application, and rapidly declining thereafter. Frozen section showed complete riboflavin penetration of the cornea immediately following pretreatment of deepithelialized corneas for all pretreatment parameters, and little penetration of corneas with intact epithelium.
Stromal riboflavin uptake with an intact epithelium was limited. All pretreatment parameters resulted in complete diffusion of riboflavin through deepithelialized corneas. Corneas pretreated for 10-30 min achieved the highest stromal riboflavin concentrations. The decrease in stromal riboflavin concentration after UVA treatment may be a result of cross-linking or binding to stroma. Therefore, there may be a need for continual riboflavin application during UVA treatment.
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